Abstract
β-Lapachone (β-lap) effectively killed MCF-7 and T47D cell lines via apoptosis in a cell-cycle-independent manner. However, the mechanism by which this compound activated downstream proteolytic execution processes were studied. At low concentrations, β-lap activated the caspase-mediated pathway, similar to the topoisomerase I poison, topotecan; apoptotic reactions caused by both agents at these doses were inhibited by zVAD-fmk. However at higher doses of β-lap, a novel non-caspase-mediated 'atypical' cleavage of PARP (i.e., an ~60-kDa cleavage fragment) was observed. Atypical PARP cleavage directly correlated with apoptosis in MCF-7 cells and was inhibited by the global cysteine protease inhibitors iodoacetamide and N- ethylmaleimide. This cleavage was insensitive to inhibitors of caspases, granzyme B, cathepsins B and L, trypsin, and chymotrypsin-like proteases. The protease responsible appears to be calcium-dependent and the concomitant cleavage of PARP and p53 was consistent with a β-lap-mediated activation of calpain. β-Lap exposure also stimulated the cleavage of lamin B, a putative caspase 6 substrate. Reexpression of procaspase-3 into caspase-3-null MCF-7 cells did not affect this atypical PARP proteolytic pathway. These findings demonstrate that β-lap kills cells through the cell-cycle-independent activation of a noncaspase proteolytic pathway. (C) 2000 Academic Press.
Original language | English (US) |
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Pages (from-to) | 144-155 |
Number of pages | 12 |
Journal | Experimental Cell Research |
Volume | 255 |
Issue number | 2 |
DOIs | |
State | Published - Mar 15 2000 |
Keywords
- Apoptosis
- Breast cancer
- Calpain
- Caspase
- PARP
- Poly(ADP)-ribose polymerase
- Topotecan
- β-lapachone
ASJC Scopus subject areas
- Cell Biology